BACKGROUND. RNAi is a powerful genetic tool for dissecting functional vulnerabilities in cancer cells. Current generation of siRNA libraries are not knockdown validated and therefore do not provide full saturation. PURPOSE. In this project we aim to accomplish the following goals: 1) constructing a high quality siRNA library targeting Ras family and related genes in which individual siRNAs is knockdown validated; 2) use this siRNA library to screen a panel of human cancer cell lines with or without KRAS mutations to identify those siRNAs that show synthetic lethality among the KRAS mutant cell lines; and 3) functional dissection of the mechanisms of candidate synthetic lethal genes using molecular and pharmacological tools. SIGNIFICANT MATERIALS AND METHODS. Knockdown validated siRNA library targeting Ras Ras pathway genes. ACCOMPLISHMENT. We have screened a large number of siRNAs against Ras pathway genes and identified those that gave potent knockdown. We have demonstrated that potent siRNAs can be combined to target multiple genes and in collaboration we showed that these siRNAs can be delivered in vivo to target undruggable targets including KRAS. The first phase of this study has been published.